Mitochondrial DNA (mtDNA) disorders encompass a broad group of inherited neuromuscular disorders caused by a biochemical defect of the respiratory chain and oxidative phosphorylation system, with a minimum prevalence of around 1:5,000 live births. The clinical diagnosis of individuals with mitochondrial disease poses a major challenge due to the clinical heterogeneity and therefore, proper genetic diagnosis of mtDNA disorders is important for prognosis and to provide counselling. The purpose of the study is to present the current strategies for the genetic diagnosis of mtDNA disorders used by the Laboratory of Human Molecular Genetics of The Mother and Child Institute. The present study was carried out on 43 patients with the characteristic phenotype of mitochondrial diseases. The strategy for the molecular-genetic diagnosis of mitochondrial disorders caused by mtDNA mutations involved performing HRM (High Resolution Melting) analysis in order to test patients for the most common pathogenic point mutations, followed by partial sequencing of the mitochondrial genome by the Sanger technique. After testing 43 patients by the HRM technique, 3 patients with pathogenic mutations associated with mitochondrial pathologies were identified (7%). The Sanger sequencing technique was performed in 23 patients, and as a result, pathogenic or potentially pathogenic mutations associated with mitochondrial pathology were identified in 9 patients (39%). The molecular-genetic techniques available in the Republic of Moldova can effectively be used for the diagnosis of mitochondrial diseases caused by mtDNA mutations. The current strategies for the genetic diagnosis of mtDNA disorders used by the research team within the Human Molecular Genetics Laboratory of the Institute of Mother and Child, allow establishing in a short period of time, but also cost effectively, the presence or absence of pathogenic variants at the level of mtDNA. In the group of 43 investigated patients, it was possible to identify pathogenic and potentially pathogenic variants associated with the patients' phenotype in 28%.